Project summary Osteoarthritis (OA) is a major cause of morbidity in our aging population. Cartilage destruction in OA is an active process, driven by expression of catabolic genes by articular chondrocytes. In normal endochondral bone development, parathyroid hormone related protein (PTHrP) inhibits expression of a similar catabolic gene profile in growth plate chondrocytes. Therefore, treatments that mimic PTHrP action represent promising novel disease-modifying treatment strategies for osteoarthritis. Recent mechanistic studies examining the intracellular signaling cascade evoked by parathyroid hormone (PTH) and PTHrP in bone cells have pointed towards an important role for salt inducible kinases (SIKs). PTH/PTHrP signaling inhibits cellular SIK activity. Therefore, orally-available small molecule SIK inhibitors, like YKL-05-099, mimic PTH/PTHrP action in bone, both in vitro and in vivo. Aim 1 of this proposal will assess the efficacy of YKL-05-099 in mice subjected to post-surgical osteoarthritis via destabilization of the medial meniscus (DMM). Mice will be subjected to DMM surgery, then treated with vehicle or YKL-05-099 for 6 or 14 weeks. Severity of cartilage destruction will be assessed by multiple techniques including 3D rapid confocal imaging, OARSI scoring, and quantitative histomorphometry. Complementing these phenotypic studies, molecular markers of cartilage destruction will be assessed in parallel. In Aim 2 of this proposal, we will establish a robust model to culture epiphyseal chondrocytes in order to dissect the mechanisms through which PTHrP and SIK3 control chondrocyte biology. Following published protocols, epiphyseal chondrocytes will be isolated and cells obtained characterized using a multiple approaches including targeted gene expression profiling and Alcian Blue staining. Then, responsiveness to PTHrP and/or YKL-05-099 will be assessed using cAMP radioimmunoassays, targeted gene expression profiling, and phospho-specific immunoblotting. Taken together, these studies will establish the efficacy of YKL-05-099 in a murine model of post- surgical osteoarthritis, and set the stage for us to determine how the PTHrP/SIK signaling axis controls chondrocyte biology.